MicroRNA phenotypic screening accomplished using libraries of microRNA mimics may allow the identification and validation of microRNA function in both the pathology of disease and the profiling of responses to therapeutic intervention. This project is focused on the concept that by comparing the results of microRNA screens from cells that express wild-type KRAS and mutant KRAS, we can not only identify microRNAs involved in context-dependent survival, but also genes and pathways that would be obscured by due to functional redundancy in a conventional siRNA screen. We will focus our attention on microRNAs that influence the viability of lung cancer cells. Lung cancer, especially non-small cell lung cancer (NSCLC), is currently the leading cause of fatalities in cancer patients in the United States. Very few effective treatment options are available for NSCLC, and many vary dramatically in efficacy depending on the genetic status of the tumor. In particular, the presence of mutant KRAS in NSCLC serves as a negative predictor for drug efficacy. KRAS mutations are found in a significant proportion of NSCLC tumors, and tumors harboring these mutations are resistant to treatment. Using microRNA mimic library screens we plan to identify synthetic lethal microRNAs in the background of KRAS-dependence. Employing the comprehensive library of microRNA mimics with an endpoint assay for cell viability, we can identify microRNAs that modulate the viability or paclitaxel sensitivity in KRAS-dependent NSCLC cell line NCI-H358 with minimal effect in the wild-type KRAS cell line NCI-H1993. Candidate microRNAs will be further validated in a panel of cell lines including KRAS mutant, KRAS wild-type and immortalized human bronchial epithelial cell lines. We will next investigate the mechanism(s) that contribute to the difference in survival induced by introduction of the specific candidate microRNAs in these cell lines. This approach will also enable us to identify potential microRNAs that may influence the growth and viability of lung tumors. To investigate this, we will perform xenograft studies in mice upon over-expression of candidate microRNAs in KRAS mutant and KRAS wild-type cells. To further establish the anti-cancer activity of the candidate microRNAs on NSCLC cells, we will also conduct follow-up analysis of these microRNAs in mouse models of lung cancer. The microRNAs identified in this project will allow us to elucidate vulnerable pathways in KRAS-dependent tumors that can be exploited for a therapeutic or predictive benefit in a typically difficult to manage class of tumor.